Lipopolysaccharide activates nuclear factor-KappaB through toll-like receptors and related molecules in cultured biliary epithelial cells
Lipopolysaccharide activates nuclear factor-KappaB through toll-like receptors and related molecules in cultured biliary epithelial cells
复制标题
DOI：
10.1097/01.lab.0000097190.56734.fe
复制
发表时间：
2003-11-01
影响因子：
5
通讯作者：
Nakanuma, Y
中科院分区：
文献类型：
--
作者：
Harada, K;Ohira, S;Nakanuma, Y
为了阐明肝内胆管树的先天免疫，我们研究了Toll样受体的表达和细胞内信号转导在胆管上皮细胞中的细菌成分的反应，通过使用培养的胆管上皮细胞（鼠胆管细胞和人胆管癌细胞系）。用逆转录、PCR和免疫组化方法检测Toll样受体在培养细胞中的表达。通过分析核因子（NF）-κ B激活和抑制研究，使用NF-κ B和丝裂原活化蛋白激酶和阻断抗体的抑制剂，脂多糖激活Toll样受体后的细胞内信号传导进行了检查。检测Toll样受体2、3、4和5以及相关分子（MD-2、MyD88和CD14）的mRNA，并在培养的细胞中表达其蛋白。脂多糖被证明与培养细胞的细胞表面结合。脂多糖处理诱导培养细胞中TNF-α的产生和NF-κ B的核转位，并增加NF-κ B-DNA结合。TNF-α的这种诱导被抗Toll样受体4抗体部分抑制。加入NF-κ B抑制剂MG 132可阻断脂多糖引起的核转位和NF-κ B结合增加。总之，脂多糖似乎在培养的胆管上皮细胞中形成CD14、Toll样受体4、MD-2和MyD88的受体复合物，并且似乎调节NF-κ B的活化和TNF-α的合成。使用Toll样受体和胆管上皮细胞中的相关分子识别病原体相关分子模式，这在体外研究中得到证实，可能参与体内肝内胆管树的免疫病理学。
To clarify the innate immunity of the intrahepatic biliary tree, we examined expression of Toll-like receptors and intracellular signalings in biliary epithelial cells in response to bacterial components by using cultured biliary epithelial cells (murine biliary cells and human cholangiocarcinoma cell lines). The expression of Toll-like receptors in cultured cells was examined by reverse transcription and PCR and immunohistochemistry. Intracellular signalings after Toll-like receptors activation by lipopolysaccharide was examined by analysis of nuclear factor (NF)-kappaB activation and inhibition studies using inhibitors for NF-kappaB and mitogen-activated protein kinase and blocking antibody. The mRNAs of Toll-like receptors 2, 3, 4, and 5, and related molecules (MD-2, MyD88, and CD14) were detected, and their proteins were expressed in cultured cells. Lipopolysaccharide was shown to bind to the cell surface of cultured cells. Lipopolysaccharide treatment induced the production of TNF-alpha, and nuclear translocation of NF-kappaB and increased NF-kappaB-DNA binding in cultured cells. This induction of TNF-alpha was partially inhibited by anti-Toll-like receptor 4 antibody. The nuclear translocation and increased binding of NF-kappaB by lipopolysaccharide were blocked by addition of MG132, an inhibitor of NF-kappaB. In conclusion, lipopolysaccharide appears to form a receptor complex of CD14, Toll-like receptor 4, MD-2, and MyD88 in cultured biliary epithelial cells and seems to regulate activation of NF-kappaB and synthesis of TNF-alpha. The recognition of pathogen-associated molecular patterns using Toll-like receptors and related molecules in biliary epithelial cells, which is demonstrated in this in vitro study, may participate in an immunopathology of the intrahepatic biliary tree in vivo.